Underactive bladder (UAB), characterized by a complex set of symptoms with few treatment options, can significantly reduce the quality of life of affected people. UAB is characterized by hyperplasia and fibrosis of the bladder wall as well as decreased bladder compliance. Pirfenidone is a powerful anti-fibrotic agent that inhibits the progression of fibrosis in people with idiopathic pulmonary fibrosis. In the current study, we evaluated the efficacy of pirfenidone in the treatment of bladder fibrosis in a UAB rat model. UAB was induced by crushing damage to nerve bundles in the major pelvic ganglion. Forty-two days after surgery, 1 mL distilled water containing pirfenidone (100, 300, or 500 mg/kg) was orally administered once every 2 days for a total of 10 times for 20 days to the rats in the pirfenidone-treated groups. Crushing damage to the nerve bundles caused voiding dysfunction, resulting in increased bladder weight and the level of fibrous related factors in the bladder, leading to UAB symptoms. Pirfenidone treatment improved urinary function, increased bladder weight and suppressed the expression of fibrosis factors. The results of this experiment suggest that pirfenidone can be used to ameliorate difficult-to-treat urological conditions such as bladder fibrosis. Therefore, pirfenidone treatment can be considered an option to improve voiding function in patient with incurable UAB.

The present study aimed to investigate the acute effects and the mechanism of ketamine on nicotine-induced relaxation of the corpus cavernosum (CC) in mice. This study measured the intra-cavernosal pressure (ICP) of male C57BL/6 mice and the CC muscle activities using an organ bath wire myograph. Various drugs were used to investigate the mechanism of ketamine on nicotine-induced relaxation. Direct ketamine injection into the major pelvic ganglion (MPG) inhibited MPG-induced increases in ICP. D-serine/L-glutamate-induced relaxation of the CC was inhibited by MK-801 (N-methyl-D-aspartate (NMDA) receptor inhibitor), and nicotine-induced relaxation was enhanced by D-serine/L-glutamate. NMDA had no effect on CC relaxation. Nicotine-induced relaxation of the CC was suppressed by mecamylamine (a non-selective nicotinic acetylcholine receptor antagonist), lidocaine, guanethidine (an adrenergic neuronal blocker), Nw-nitro-L-arginine (a non-selective nitric oxide synthase inhibitor), MK-801, and ketamine. This relaxation was almost completely inhibited in CC strips pretreated with 6-hydroxydopamine (a neurotoxic synthetic organic compound). Ketamine inhibited cavernosal nerve neurotransmission via direct action on the ganglion and impaired nicotine-induced CC relaxation. The relaxation of the CC was dependent on the interaction of the sympathetic and parasympathetic nerves, which may be mediated by the NMDA receptor.

Micromotion is an autonomous intramural movement of the bladder, and is believed to be an initial step in the generation of urinary urgency. Therefore, controlling micromotion may be a novel target in overactive bladder (OAB) treatment. However, developing micromotion treatment has been limited by the absence of a standardized animal model. We attempted to create a micromotion animal model and investigated the effectiveness of a β3 -adrenoceptor agonist (CL316,243) on micromotion.
Bilateral major pelvic ganglia (MPGs) were excised in 18 male Sprague-Dawley rats, resulting in an almost completely denervated bladder. On postoperative Day 7, cystometry was performed. Rats were divided into three treatment groups: CL316,243; β3- adrenoceptor antagonist (SR59230A) pretreated CL316,243; and a nonselective antimuscarinic agent (oxybutynin). Changes in micromotion were evaluated after the intra-arterial administration of each agent.
Low-amplitude oscillations in intravesical pressure (micromotion) were observed 1 week after MPGs excision. Micromotion frequency significantly (p = 0.003) decreased (2.17 ± 3.54 times/5 min) with CL316,243 compared with vehicle (6.33 ± 1.97 times/5 min). Micromotion amplitude also decreased with CL316,243 (1.15 ± 1.93 cmH2 O) compared with vehicle (5.96 ± 5.12 cmH2 O), approaching conventional significance (p = 0.090). No significant decreases in frequency or amplitude were observed with oxybutynin treatment.
Systemic administration of the β3 -adrenoceptor agonist CL316,243 effectively controlled micromotion in bilateral MPGs-excised, almost completely denervated rat bladders. This result indicates that β3 -adrenoceptor agonist may affect the bladder directly, suggesting that it might be effective for overall OAB, regardless of the presence or level of neurological deficits. Bilateral MPGs-excised rats are considered a plausible micromotion animal model suitable for future research.

To map sensory and pelvic postganglionic neurons from three different regions of the female rat urethra.
The neuronal tracer True Blue (TB) was injected into the pre-pelvic, pelvic, and clitoral regions of the urethra from female Wistar rats. Seven days after TB injection, TB+ cells from the dorsal root ganglia (DRGs) and the major pelvic ganglion (MPG) were examined. The number and morphometry of TB+ cells were determined.
TB+ cells were mainly distributed in lumbar 1 (L1), lumbar 2 (L2), lumbar 6 (L6), and sacral 1 (S1) DRGs, and in the MPG. The mean number of sensory neurons was 1200 ± 143. TB injection in pre-pelvic and pelvic urethra labeled neurons in L1, L2, L6, and S1 DRGs. TB injection in clitoral urethra labeled neurons in L6 and S1 DRGs. L6 DRG contained >50% of the total urethral TB+ neurons, and ~80% of the clitoral region. The mean value of the total number of MPG TB+ neurons was 1217 ± 72. DRG and MPG neurons projecting to the urethra presented a somatotopic distribution.
The results demonstrated that L6 DRG is the major supplier of afferent innervation to the urethra, and that the distal urethral region is exclusively innervated by lower lumbosacral DRGs. Considering that electrical stimulation of sensory pudendal nerve improves overactive bladder, and that most of the sensory neurons in the distal urethra are from L6 DRG, electrical stimulation of this ganglion may be an innovative and effective neuromodulation therapy for neurogenic urinary dysfunctions.

The primary function of the urinary bladder is to store urine (continence) until a suitable time for voiding (micturition). These distinct processes are determined by the coordinated activation of sensory and motor components of the nervous system, which matures to enable voluntary control at the time of weaning. Our aim was to define the development and maturation of the nerve-organ interface of the mouse urinary bladder by mapping the organ and tissue distribution of major classes of autonomic (motor) and sensory axons. Innervation of the bladder was evident from E13 and progressed dorsoventrally. Increasing defasciculation of axon bundles to single axons within the muscle occurred through the prenatal period, and in several classes of axons underwent further maturation until P7. Urothelial innervation occurred more slowly than muscle innervation and showed a clear regional difference, from E18 the bladder neck having the highest density of urothelial nerves. These features of innervation were similar in males and females but varied in timing and tissue density between different axon classes. We also analysed the pelvic ganglion, the major source of motor axons that innervate the lower urinary tract and other pelvic organs. Cholinergic, nitrergic (subset of cholinergic) and noradrenergic neuronal cell bodies were present prior to visualization of these axon classes within the bladder. Examination of cholinergic structures within the pelvic ganglion indicated that connections from spinal preganglionic neurons to pelvic ganglion neurons were already present by E12, a time at which these autonomic ganglion neurons had not yet innervated the bladder. These putative preganglionic inputs increased in density prior to birth as axon terminal fields continued to expand within the bladder tissues. Our studies also revealed in numerous pelvic ganglion neurons an unexpected transient expression of calcitonin gene-related peptide, a peptide commonly used to visualise the peptidergic class of visceral sensory axons. Together, our outcomes enhance our understanding of neural regulatory elements in the lower urinary tract during development and provide a foundation for studies of plasticity and regenerative capacity in the adult system.

To investigate the relationship between lower urinary tract function and the accessory nerve (ACN) arising from the major pelvic ganglion (MPG).
Ten-week-old male Wistar/ST rats were randomly divided into eight groups according to the type of treatment (sham or bilateral accessory nerve injury [BACNI]) and the duration of observation (3 days, 1 week, 2 weeks, or 4 weeks: Sham-3d, Sham-1w, Sham-2w, Sham-4w, BACNI-3d, BACNI-1w, BACNI-2ws, and BACNI-4w. BACNI was induced in the following manner: the ACN was crushed for 1 min (2 mm away from the MPG) using reverse-action tweezers. The same procedure was performed on both sides. On the last day of each observation period, the bladder function was measured by awake cystometry, and histological evaluation was performed.
All rats in the Sham groups micturated normally. In the BACNI-3d and BACNI-1w groups, all rats showed symptoms of overflow urinary incontinence (OUI). This OUI improved gradually over time. The bladder\'s size in the BACNI group was significantly larger than that in the Sham group (p < .01). In addition, fibrosis was observed in the subserosa of the bladder of rats in BACNI groups.
The BACNI model rats exhibited OUI, suggesting that ACN is involved in the lower urinary tract function. It might be possible that ACN controls the function of either the bladder, the urethra, or both.

Neurogenic erectile dysfunction (ED) is often refractory to treatment because of insufficient functional nerve recovery after injury or insult. Noninvasive mechano-biological intervention, such as microenergy acoustic pulse (MAP), low-intensity pulsed ultrasound, and low-intensity extracorporeal shockwave treatment, is an optimal approach to stimulate nerve regeneration.
To establish a new model in vitro to simulate nerve injury in neurogenic ED and to explore the mechanisms of MAP in vitro.
Sprague-Dawley rats were used to isolate Schwann cells (SCs), major pelvic ganglion (MPG), and cavernous nerve with MPG (CN/MPG). SCs were then treated with MAP (0.033 mJ/mm2, 1 Hz, 100 pulses), and SC exosomes were isolated. The MPG and CN/MPG were treated with MAP (0.033 mJ/mm2, 1 Hz) at different dosages (25, 50, 100, 200, or 300 pulses) or exosomes derived from MAP-treated SCs in vitro.
Neurite growth from the MPG fragments and CN was photographed and measured. Expression of neurotropic factors (brain-derived neurotrophic factor, nerve growth factor, and neurotrophin-3) was checked.
Neurite outgrowth from MPG and CN/MPG was enhanced by MAP in a dosage response manner, peaking at 100 pulses. MAP promoted SC proliferation, neurotropic factor (brain-derived neurotrophic factor, nerve growth factor, and neurotrophin-3) expression, and exosome secretion. SC-derived exosomes significantly enhanced neurite outgrowth from MPG in vitro.
MAP may have utility in the treatment of neurogenic ED by SC-derived exosomes.
We confirmed that MAP enhances penile nerve regeneration through exsomes. Limitations of this study include that our study did not explore the exact mechanisms of how MAP increases SC exosome secretion nor whether MAP modulates the content of exosomes.
This study revealed that neurite outgrowth from MPG was enhanced by MAP and by SC-derived exosomes which were isolated after MAP treatment. Our findings indicate that one mechanism by which MAP induces nerve regeneration is by stimulation of SCs to secrete exosomes. Peng D, Reed-Maldonado AB, Zhou F, et al. Exosome Released From Schwann Cells May Be Involved in Microenergy Acoustic Pulse-Associated Cavernous Nerve Regeneration. J Sex Med 2020;17:1618-1628.

The aims of the present study were to describe, in male rats, the anatomical organization of the major and accessory pelvic ganglia (MPG, AG; respectively), the interrelationship of the pelvic plexus components, and the morphometry of the pelvic postganglionic neurons. Anatomical, histochemical and histological studies were performed in anesthetized adult Wistar male rats. We found that the pelvic plexus consists of intricate neural circuits composed of two MPG, and three pairs of AG (AGI, AGII, AGIII) anatomically interrelated through ipsilateral and contralateral commissural nerves. Around 30 nerves emerge from each MPG and 17 from AGI and AGII. The MPG efferent nerves spread out preganglionic information to several pelvic organs controlling urinary, bowel, reproductive and sexual functions, while AG innervation is more regional, and it is confined to reproductive organs located in the rostral region of the urogenital tract. Both MPG and AG contain nerve fascicles, blood vessels, small intensely fluorescent cells, satellite cells and oval neuronal somata with one to three nucleoli. The soma area of AG neurons is larger than those of MPG neurons (p < 0.005). The MPG contains about 75% of the total pelvic postganglionic neurons. Our findings corroborated previous reports about MPG inputs, and add new information regarding pelvic ganglia efferent branches, AG neurons (number and morphometry), and neural interrelationship between the pelvic plexus components. This information will be useful in designing future studies about the role of pelvic innervation in the physiology and pathophysiology of pelvic functions.

Neurogenic erectile dysfunction is a common sequela of radical prostatectomy. The etiology involves injury to the autonomic cavernous nerves, which arise from the major pelvic ganglion (MPG), and subsequent neuroinflammation, which leads to recruitment of macrophages to the injury site. Currently, two macrophage phenotypes are known: neurotoxic M1 macrophages and neuroprotective M2 macrophages.
To examine whether bilateral cavernous nerve injury (BCNI) in a rat model of erectile dysfunction would increase recruitment of neurotoxic M1 macrophages to the MPG.
Male Sprague-Dawley rats underwent BCNI and the MPG was harvested at various time points after injury. The corpora cavernosa was used to evaluate tissue myographic responses to electrical field stimulation ex vivo. Quantitative real-time polymerase chain reaction was used to examine the gene expression of global macrophage markers, M1 macrophage markers, M2 macrophage markers, and cytokines and chemokines in the MPG. Mathematical calculation of the M1/M2 index was used to quantify macrophage changes temporally. Western blot of MPG tissues was used to evaluate the protein amount of M1 and M2 macrophage markers quantitatively. Immunohistochemistry staining of MPGs for CD68, CD86, and CD206 was used to characterize M1 and M2 macrophage infiltration.
Corpora cavernosa responsiveness ex vivo; gene (quantitative real-time polymerase chain reaction) and protein (western blot) expressions of M1 and M2 markers, cytokines, and chemokines; and immunohistochemical localization of M1 and M2 macrophages.
BCNI impaired the corporal parasympathetic-mediated relaxation response to electrical field stimulation and enhanced the contraction response to electrical field stimulation. Gene expression of proinflammatory (Il1b, Il16, Tnfa, Tgfb, Ccl2, Ccr2) and anti-inflammatory (Il10) cytokines was upregulated in the MPG 48 hours after injury. M1 markers (CD86, inducible nitric oxide synthase, interleukin-1β) and M2 markers (CD206, arginase-1, interleukin-10) were increased after BCNI in the MPG, with the M1/M2 index above 1.0 indicating that more M1 than M2 macrophages were recruited to the MPG. Protein expression of the M1 macrophage marker (inducible nitric oxide synthase) was increased in MPGs after BCNI. However, the protein amount of M2 macrophage markers (arginase-1) remained unchanged. Immunohistochemical characterization demonstrated predominant increases in M1 (CD68+CD86+) macrophages in the MPG after BCNI.
These results suggest that an increase in M1 macrophage infiltration of the MPG after BCNI is associated with impaired neurogenically mediated erectile tissue physiology ex vivo and thus has significant implications for cavernous nerve axonal repair. Future studies are needed to demonstrate that inhibition of M1 macrophage recruitment prevents erectile dysfunction after CNI.

The aim of the present study was to use a model of simulated human childbirth in rats to determine the damage to genitourinary structures and behavioral signs of urinary dysfunction induced by vaginal distension (VD) in female rats. In experiment 1, the length of the genitourinary tract and the nerves associated with it were measured immediately after simulated human delivery induced by VD or sham (SH) procedures. Electroneurograms of the dorsal nerve of the clitoris (DNC) were also recorded. In experiment 2, histological characteristics of the bladder and major pelvic ganglion of VD and SH rats were evaluated. In experiment 3, urinary parameters were determined in conscious animals during 6 h of dark and 6 h of light before and 3 days after VD or SH procedures. VD significantly increased distal vagina width (P < 0.001) and the length of the motor branch of the sacral plexus (P < 0.05), DNC (P < 0.05), and vesical nerves (P < 0.01) and decreased DNC frequency and amplitude of firing. VD occluded the pelvic urethra, inducing urinary retention, hematomas in the bladder, and thinness of the epithelial (P < 0.05) and detrusor (P < 0.01) layers of the bladder. Major pelvic ganglion parameters were not modified after VD. Rats dripped urine in unusual places to void, without the stereotyped behavior of micturition after VD. The neuroanatomic injuries after VD occur alongside behavioral signs of urinary incontinence as determined by a new behavioral tool for assessing micturition in conscious animals.